1. Field of the Invention
The present invention is directed to the provision of compounds having potent calcium antagonist and antioxidant activity, and to the use of those compounds as cellular protective agents. The invention is further directed to the provision of methods for synthesizing the compounds of the invention and to compounds formed as intermediates during the synthesis. The invention is particularly directed to the use of the compounds of the present invention to prevent or reduce cellular damage associated with ophthalmic diseases or injuries.
2. Discussion of Related Art
In a biological system under stress induced by trauma, ischemia-reperfusion, depletion of natural defenses, inflammation, light damage (especially laser or intense operating room light), or degenerative conditions, damage occurs which can result in an increase in cellular free calcium and/or an increase in oxidative damage. Both these changes are components of the common pathway of cell death. The result of these changes is the initiation of a cascade of cellular destruction, loss of cellular function and ultimately cell loss. The loss of critical cellular components can result in organ damage and loss of organ function. Loss of function can be caused by an acute insult or may be the result of the cumulative effects of chronic insult. The following texts may be referred to for further details concerning these phenomena:
Prog. Neuro-Psychopharmacol. and Biol. Pysch., volume 17, pages 21-70 (1993); PA1 Age, volume 16, pages 23-30 (1993); PA1 Chem. Res. Tox., volume 32, pages 2-18 (1993); and PA1 Ann. Neurol., volume 32, pages S33-42 (1992). PA1 Med. Res. Review, volume 9, pages 123-80 (1989); PA1 Pharmacol. Review, volume 38(4), pages 321-416 (1986); PA1 Cardiovasc. Drugs and Therapy, volume 6, pages 35-39 (1992); PA1 Science, volume 235, pages 46-52 (1987); PA1 Chem.-Biol. Interactions, pages 1-23 (1991); and PA1 Biochemical Pharmacol., volume 43(1), pages 39-46 (1992). PA1 Arch. Pharmacol., volume 325, pages 129-146 (1992); PA1 Free Rad. Biol. Med., volume 6, pages 209-224; PA1 Free Rad. Biol. Med., volume 11, pages 215-232 (1991); PA1 Eur. J. Pharmacol., volume 210, pages 85-90 (1992); PA1 J. Photochem., Photobiol. Biol., volume 8, pages 211-224 (1991); PA1 Pharmacol. and Tox., volume 70, pages 271-277 (1992); and PA1 Medicinal Res. Rev., volume 13(2), pages 161-182 (1993). PA1 Arch. int. Pharmacodyn., volume 272, pages 283-295 (1984); PA1 Eur. J. Pharmacol., volume 204, pages 315-322 (1991); and PA1 Meth. and Find Exp. Clin. Pharmacol., volume 11(10), pages 607-612 (1989). PA1 Free Rad. Biol. and Med., volume 12, pages 183-187 (1992); PA1 Res. Commun. in Chem. Path. and Pharmacol., volume 76(3), pages 367-370 (1992); PA1 J. Mol. Cell Cardiol., volume 22, pages 1199-1208 (1990); PA1 Circulation Res., volume 66(5), pages 1449-1452 (1990); PA1 J. Cardiovas. Pharmacol., volume 18(Suppl. 1) pages S6-S10 (1991); PA1 Basic Res. in Cardiology, volume 87, pages 148-160 (1992); PA1 Free Rad. Res. Comms., volume 15(2), pages 91-100 (1991); and PA1 Biochem. Pharmacol., volume 37(21), page 4197 (1988). PA1 A is an antioxidant; PA1 Y is (CH.sub.2).sub.n or CH.dbd.CH(CH.sub.2).sub.n, wherein n is 1 to 6; and PA1 B is selected from the following groups: ##STR1## wherein: n' is 1 to 6; PA1 Z is H, CN or OH; PA1 X is F, Cl, I, Br, OH, OR', SH, S(O).sub.m R', CN or NO.sub.2, wherein R' is branched PA1 or unbranched C.sub.1 to C.sub.6 alkyl and m is 0, 1 or 2; and PA1 o is 0 to 3. PA1 1) provide greater than 20% quench of the free radical at concentrations of DPPH and the test agent equal to 10.sup.-4 M, in accordance with the above-cited DPPH assay; PA1 2) demonstrate an IC.sub.50 of less than 20 .mu.M, in accordance with the above cited liposome assay; or PA1 3) demonstrate an IC.sub.50 of less than 20 .mu.M, in accordance with the above-cited liver microsome assay. PA1 a) preventing the flux through voltage-sensitive calcium channels (N,L,T,P); PA1 b) blocking flux through receptor operated calcium channels; PA1 c) preventing the release of calcium sequestered in sarcoplasmic reticulum; or PA1 d) blocking nonspecific channels (i.e., reversing sodium/calcium exchangers or blocking calcium flux through a sodium channel). PA1 1) radioligand binding assays, wherein radiolabeled nitrendipine is displaced from rat brain cortices (minimum activity: IC.sub.50 of less than 20 .mu.M), as described in Life Science, volume 30, pages 2191-2202 (1979) and Procedures of the National Academy of Science, USA, volume 79, pages 3656-3650 (1982); PA1 2) calcium antagonist binding assays, such as the relaxation of pre-contracted rabbit aortic strips of greater than 7.0, as described in Journal of Medicinal Chemistry, volume 34, pages 3011-3022 (1991) and references cited therein (minimum activity: IC.sub.50 value less than 20 .mu.M); PA1 3) inhibition of calcium flux in a cellular system, as measured by a fluorescent dye, in accordance with the procedures described in Journal of Cardiovascular Pharmacology, volume 17, pages 41-53 (1991), and references cited therein, (minimum activity: IC.sub.50 of less than 100 nm); or PA1 4) inhibition of calcium induced contractions of rabbit thoracic aortic strips, in accordance with the procedures described in Journal Cardiovascular Pharmacology, volume 17, pages 41-53 (1991), and references cited therein (minimum activity: pA.sub.2 greater than 7).
Calcium flux is a necessary part of normal cell function. The level of intracellular free calcium is highly regulated. Both receptor-operated and voltage-sensitive channels control cell signaling and stimulus response. Multiple voltage-sensitive calcium channels have been identified. These include the N, T, P, and L channels. The following publications may be referred to for further background concerning the regulation of intracellular free calcium levels:
Over-stimulation of the cell or cellular system or the defective regulation of intracellular free calcium can result in increased intracellular free calcium levels. This can lead to the initiation of a chain of biochemical processes which can lead to cell death. Agents that modulate increases in intracellular free calcium concentration can moderate the deleterious effects of over-stimulation or defective regulation. See PNAS, volume 89, pages 435-39 (1992), and references cited above. In addition, a compound that acts as a calcium antagonist can provide an additional beneficial effect by improving blood flow, reducing ischemic insult and facilitating repair. See Naunyn-Schmiedeberg's Acta Pharmacol., volume 335, pages 680-685 (1987). As utilized herein, the term "calcium antagonists" refers to organic molecules which inhibit increases in intracellular free calcium concentrations.
Agents that act as antioxidants can protect against oxidative damage associated with cellular stress. Such protection has been the subject of numerous scientific publications, including the following:
The combined use of two or more compounds having calcium antagonist and antioxidant activity, respectively, is discussed in Experimental Eye Research, volume 5, pages 71-78 (1993). The provision of compounds having both calcium antagonist and antioxidant activity is discussed in the following patent publications: EP 267 155A and WO 89/05803 A1.
One compound known to have calcium antagonist activity, flunarizine, has also been reported to have free radical scavenging activity. See:
In addition, other classes of calcium antagonists have been reported to have antioxidant activity. See:
However, in most cases the antioxidant effect reported is weak and not clinically relevant. This is pointed out in Biochem. Pharmacol., volume 42(4), pages 735-743 (1991), and Biochem. Phannacol., 38(20), pages 3601-3610 (1989). In addition, it is believed that a number of the effects attributed to the free radical scavenging effect of flunarizine might actually be an effect of its calcium antagonist activity since this activity was poorly understood in the early 1980's.
The present invention is directed to the provision of new compounds that have both potent calcium antagonist and potent antioxidant activity in a single molecule. The use of a single chemical entity with potent antioxidant and potent calcium antagonist activity provides increased protection relative to the use of a compound with singular activity. The advantage of a single agent with both activities over a combination of two components would be realized by the uniform delivery of an active molecule simplifying issues of drug metabolism and delivery.